CN104897907B - A kind of test kit detecting glycolated hemoglobin and detection method thereof - Google Patents
A kind of test kit detecting glycolated hemoglobin and detection method thereof Download PDFInfo
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- CN104897907B CN104897907B CN201510262100.6A CN201510262100A CN104897907B CN 104897907 B CN104897907 B CN 104897907B CN 201510262100 A CN201510262100 A CN 201510262100A CN 104897907 B CN104897907 B CN 104897907B
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/72—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood pigments, e.g. haemoglobin, bilirubin or other porphyrins; involving occult blood
- G01N33/721—Haemoglobin
- G01N33/723—Glycosylated haemoglobin
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/72—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving blood pigments, e.g. haemoglobin, bilirubin or other porphyrins; involving occult blood
- G01N33/721—Haemoglobin
- G01N33/726—Devices
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/795—Porphyrin- or corrin-ring-containing peptides
- G01N2333/805—Haemoglobins; Myoglobins
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/04—Endocrine or metabolic disorders
- G01N2800/042—Disorders of carbohydrate metabolism, e.g. diabetes, glucose metabolism
Abstract
The present invention relates to glycolated hemoglobin detection technique field, it is specifically related to a kind of test kit detecting glycolated hemoglobin and detection method thereof, described test kit includes: the first reagent, and this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, the borate derivative of precipitation total hemoglobin；Second reagent, this second reagent includes the buffer of the borate derivative for washing away uncombined glycolated hemoglobin；And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.The test kit of the present invention have only to trace whole blood live peripheral blood sample, the content of detection by quantitative glycolated hemoglobin can be realized in 23 minutes, greatly improve the speed of examination, have highly sensitive, specificity good and advantage easy and simple to handle；Preparation method is simple, it is easy to large-scale production.
Description
Technical field
The present invention relates to glycolated hemoglobin detection technique field, be specifically related to a kind of reagent detecting glycolated hemoglobin
Box and detection method thereof.
Background technology
The incretion metabolism disease that diabetes are one group of cause of disease and pathogenesis is not yet fully apparent from, current sickness rate is the most secondary
In cardiovascular disease and tumor.Recent years, the sickness rate of diabetes was continuous ascendant trend, was serious threat human health
Worldwide public health problem.Traditional diabetes diagnosis and Treatment monitoring use fasting glucose, post-prandial glycemia and oral Fructus Vitis viniferae
Carbohydrate tolerance test etc., but moment blood sugar level when glycemic parameters only represents blood drawing.Nearly car comes, glycolated hemoglobin
(HbA1C) detection is increasingly subject to the great attention of clinic.Glycolated hemoglobin (HbA1C) refers in blood and glucose combines
That a part of hemoglobin.The amino of the valine of hemoglobin β-chain N end and the reversible condensation of free aldehyde radical of glucose
For aldimine (Schiff's base), then there is Amadori rearrangement reaction in aldimine, forms relatively stable N-terminal fructosyl structure
And syn diol structure unit.When the concentration of glucose in blood is higher, the saccharification hemoglobin content that human body is formed also can
Higher.In human body, the erythrocytic life-span is generally 120 days, before erythrocyte death, and glycolated hemoglobin (HbA1C) in blood
Content also remains relatively unchanged over.Glycolated hemoglobin (HbA1C) horizontal reverse has answered the average blood glucose levels in first 120 days of detection,
And whether on an empty stomach and whether use the factors such as insulin unrelated with taking blood time, patient, and therefore, glycolated hemoglobin (HbA1C)
It is the goldstandard of reaction long-term blood glucose level, is also that diabetes assist the important indicator diagnosing, monitoring, treat.
The assay method of glycolated hemoglobin (HbA1C) is varied, can be divided into two big classes on the whole: a class be based on
The electric charge of HbA1C with Hb is different, such as ion exchange chromatography, electrophoresis method；Another kind of is special based on the structure of saccharifying group on Hb
Point, such as affinity chromatography, immunization and enzyme process etc..
Multiple researchs show that the difference in principle and methodology determines the quality of detectable performance, and diabetics
Therapeutic goal requires that measured value is not affected by assay method, and the different GHb assay method of the most under lab application is produced
Result comparison extremely important.
Ion exchange chromatography: mainly have high performance liquid chromatography (HPLC) and manual microtrabeculae method.The method is based on blood
After hemoglobin beta chain N-terminal valine saccharifying institute electrically charged difference and set up.But the expensive equipment used due to the method, difficult
To popularize in the hospital of relatively basic unit and laboratory；Microtrabeculae method manual steps is loaded down with trivial details, and the quality of chromatography time and microtrabeculae is not
Easy to control, it is easily generated operating technology error, repeatability is not good enough；And interference factor is a lot, especially to pH value and the change of temperature
Result is disturbed by sensitivity, HbF and variation hemoglobin (HbS, HbC, HbE etc.), depending on annoyance level is according to the separating power of post,
So collection of illustrative plates must be examined.
Electrophoresis method: as a example by agargel electrophoresis, Hb under the conditions of acidic buffer (pH6.0) on agarose gel
Electrophoretic migration depends on Hb absorption situation on gel and the electric charge carried thereof.The method specimen consumption is few, and resolution is high, weight
Renaturation is good, studies have found that blood glucose value and HbAlc value have significant correlation, and result is not by temperature and the shadow of fetal hemoglobin
Ring.It addition, because its measurable Hb range of linearity wider (13.0～39.0g/L), it is possible to find abnormal Hb.The shortcoming of the method
Being to measure to be both needed to carry out in batch sample analysis, speed is slow, it is impossible to carrying out the most individual detection, automaticity is relatively every time
Difference, measured result is relevant with technical staff's scanning and the crest judgement to electrophoresis, is affected by subjective factors, and somewhat expensive,
Therefore it is not appropriate for clinical laboratory's routine to use.
Affinity chromatography: boric acid has makees what Reversible binding reacted with the cis-position glycol-based being incorporated into glucose on Hb molecule
Character.Normally used is m-aminobenzene boric acid agarose, and after blood sample is added to chromatographic column, all of GHb is combined with boric acid
Staying in post, non-GHb flows directly out chromatographic column；Add high concentration and also comprise the polyhydroxy complex of cis-position glycol-based (such as mountain
Pears alcohol), the combination of GHb and boric acid is replaced and is eluted, and measures two components, and ratio calculated respectively.Affinity chromatography
Insensitive relative to additive method with the impact of pathology hemoglobin on variation hemoglobin, but mensuration is HbA1 i.e. GHb total amount.
It addition, Gold standard is closely related with boric acid affinity chromatography, operate the most simple and easy to do, quick and precisely, stable reagent.It is reported,
This method disturbing by any haemoglobin variant in addition to Hbs and Hbc and catabolite, reliable results, be relatively suitable for
Detect at any time in clinic.
Immunoturbidimetry: utilize antigen, the principle of antibody response is measured.The β chain N-terminal of GHb provides an appearance
Easily by the epitope of antibody recognition, can be with monoclonal antibody or polyclonal antibody, the β chain N-terminal of specific recognition GHb is last
The epitope of 4～6 aminoacid compositions, in conjunction with colorimetric or turbidimetry, with GHb as standard, measures the content of HbA1c, then surveys
Determine the content of Hb, finally calculate HbA1c and account for the percentage composition of total Hb.This type of method is only as judging diabetes glucose water
Flat index, is not useable for the research of variation hemoglobin.In contrast, the method for immunoturbidimetry detection is the simplest, no
Need additionally to add instrument, provide a kind of quick, accurate, reliable, easy conventional method for clinic, have in clinical practice
The most wide prospect.
Enzyme process: Hb enzymolysis, digestion, after haemolysis processes, is become fructosyl amino acid with differential protein restriction endonuclease by whole blood, then through fruit
Produce in hydrogen peroxide (hydrogen peroxide, H2O2), the concentration of H2O2 and blood under glycoprotein amino acid oxidase effect
The content of GHb is directly proportional, and H2O2 couples with corresponding chromogen under the effect of peroxidase, thus according to color intensity of variation
H2O2 concentration can be obtained, and then learn the content of GHb in sample；Measure total Hb concentration of same pipe Digestive system simultaneously, calculate GHb and
The concentration proportion of Hb, is GHb result.This method provides a response system the most homogeneous as clinical biochemical reacts
(such as glucose, alanine aminotransferase), has good precision, can detect GHb and Hb simultaneously, and with conventional H PLC method
Good dependency is had with immunoassay.
Ion capture: application antigen antibody reaction principle, in parallel with fluorescent marker, by connect electronegative many cloudy from
Sub-complex, be adsorbed onto positively charged fiber surface, after the steps such as a series of thorough cleanings, measure fluorescence intensity change
Rate, calculates GHb concentration.Its detecting system is prone to specification and repetition, can reduce operating technology error, the sensitivity of detection and specificity
Height, batch in, interassay coefficient of variation little.This method influence factor is few to have document to report, accuracy is high, and the response rate reaches 98.85%, intersects dirty
Dye rate < 0.01%.The method is the new method grown up in recent years, uses automatic analyzer, it is adaptable to the detection of specimen in batches.
Several method in sum, otherwise reagent, instrument cost are high, and operation is complicated or accuracy is low, stable
Property is poor, and is all not suitable for community and basic hospital.Accordingly, it would be desirable to the method for existing mensuration glycolated hemoglobin ratio is carried out
Improve, propose a kind of new method and solve these problems.
Summary of the invention
In order to overcome shortcoming and defect present in prior art, it is an object of the invention to provide a kind of detection saccharifying blood
The test kit of Lactoferrin, this test kit has only to the whole blood peripheral blood sample alive of trace, can realize quantitatively inspection in 2-3 minute
Survey the content of glycolated hemoglobin, greatly improve the speed of examination, have highly sensitive, specificity good and easy and simple to handle excellent
Point；Preparation method is simple, it is easy to large-scale production.
Another object of the present invention is to provide a kind of detection method detecting glycolated hemoglobin, this detection method only needs
Want the whole blood peripheral blood sample alive of trace, the content of detection by quantitative glycolated hemoglobin can be realized in 2-3 minute, greatly carry
The high speed of examination, have highly sensitive, specificity good and advantage easy and simple to handle.
The purpose of the present invention is achieved through the following technical solutions: a kind of test kit detecting glycolated hemoglobin, described examination
Agent box includes:
First reagent, this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, precipitation total hemoglobin
Borate derivative；
Second reagent, this second reagent includes the buffer of the borate derivative for washing away uncombined glycolated hemoglobin；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
Preferably, described test kit also includes brown centrifuge tube, reagent bottle and aluminium foil bag, and brown centrifuge tube is equipped with the first examination
Agent, and brown centrifuge tube is contained in aluminium foil bag, reagent bottle is equipped with the second reagent.
Being more highly preferred to, described test kit also includes packing box, liner and capillary blood taking tube, and liner is arranged at packing box
In, capillary blood taking tube is arranged at interior lining.Chromatography device also includes cabinet, and reaction film is fixed in the inside of cabinet
Between, cabinet and reaction film are assembled into chromatography device, and the aperture of reaction film is 0.5 μm.
First reagent is borate derivative, pH8.0, total amount 200 μ L, is placed in the brown centrifuge tube of 1mL capacity；Second
Reagent is buffer, PH8.0, total amount 2mL, is placed in 5mL capacity plastic reagent bottle.
The test kit of the present invention has only to the whole blood peripheral blood sample alive of trace, can realize quantitatively inspection in 2-3 minute
Survey the content of glycolated hemoglobin, greatly improve the speed of examination, have highly sensitive, specificity good and easy and simple to handle excellent
Point；Preparation method is simple, it is easy to large-scale production.
Preferably, every liter of described first reagent includes following component:
Borate derivative 0.4-0.6g
Magnesium chloride 1.9-2.9g
Potassium chloride 22-26g
Barium chloride 6.2-8.2g
Glycyl amide hydrochloride 6.6-8.6g
Methanamide 25-35g
Potassium azide 8-12g
NPE 15-25g
Water surplus.
Preferably, the structural formula of described borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 1-6 for carbon number.
Preferably, every liter of described second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 23-33g
Potassium azide 4-6g
Sodium chloride 0.4-0.8g
Methanamide 8-12g
Triton X-100 4-6g
Water surplus.
Another object of the present invention is achieved through the following technical solutions: a kind of described above for the use of non-treatment purpose
The detection method of test kit detection glycolated hemoglobin, comprises the steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, on
Under overturn 8-12 time, fully mix, stand 1-3min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 8-12s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 8-12s, chromatography device is placed on glycolated hemoglobin analysis and detects.
Blood sample requires: the blood sample that anticoagulant blood-collecting pipe gathers, and can preserve 7 days at 2 ~ 8 DEG C；Or finger tip blood.
The Cleaning Principle of the present invention is: 1, utilize the first reagent splitting erythrocyte of special formulation, discharges HbAle egg
In vain, precipitation total hemoglobin；Meanwhile, in the first reagent in the blue borate derivative of ad hoc structure and glycolated hemoglobin
" c/s-diol " structural specificity reacts, and generates blue " borate-glycolated hemoglobin complex "；
2, take a certain amount of reactant mixture and join chromatography device, the hemoglobin of all precipitations and and glycolated hemoglobin
In conjunction with or unconjugated boric acid conjugate remain in chromatography device in the middle of reaction film on；
3, a certain amount of second reagent dropwise is to chromatography device, and the borate derivative of any uncombined glycolated hemoglobin is removed；
4, chromatography device is placed in glycolated hemoglobin analysis detection, by instrument the most quickly detection and analysis indigo plant
Color (glycolated hemoglobin) and the color intensity of red (total hemoglobin), thus measure glycolated hemoglobin in blood sample
Percentage ratio.
The detection method of the present invention has only to the whole blood peripheral blood sample alive of trace, can realize detection by quantitative in 2-3 minute
The content of glycolated hemoglobin, greatly improves the speed of examination, have highly sensitive, specificity good and advantage easy and simple to handle.
Preferably, in described step A, every liter of first reagent includes following component:
Borate derivative 0.4-0.6g
Magnesium chloride 1.9-2.9g
Potassium chloride 22-26g
Barium chloride 6.2-8.2g
Glycyl amide hydrochloride 6.6-8.6g
Methanamide 25-35g
Potassium azide 8-12g
NPE 15-25g
Water surplus.
Preferably, in described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 1-6 for carbon number.
Preferably, in described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 23-33g
Potassium azide 4-6g
Sodium chloride 0.4-0.8g
Methanamide 8-12g
Triton X-100 4-6g
Water surplus.
The beneficial effects of the present invention is: the test kit of the present invention has only to the whole blood peripheral blood sample alive of trace,
In 2-3 minute, realize the content of detection by quantitative glycolated hemoglobin, greatly improve the speed of examination, have highly sensitive,
The advantage that specificity is good He easy and simple to handle；Preparation method is simple, it is easy to large-scale production.
The detection method of the present invention has only to the whole blood peripheral blood sample alive of trace, can realize quantitatively in 2-3 minute
The content of detection glycolated hemoglobin, greatly improves the speed of examination, have highly sensitive, specificity good and easy and simple to handle
Advantage.
Present invention have the advantage that
1, adapted to instrument platform is small and exquisite, it is simple to carry；
2, instrument cost, reagent cost are far below the detection platform of other principles, are suitable for and community and township hospital；
3, easy and simple to handle, detection is rapidly (each pattern detection time ＜ 3 minutes)；
4, result is accurate, stable, and interference factor is few；
5, required sample size is few, detection of bleeding (sample size 5 μ L).
Detailed description of the invention
For the ease of the understanding of those skilled in the art, below in conjunction with embodiment, the present invention is further illustrated, real
The content that the mode of executing is mentioned not limitation of the invention.
Embodiment 1
A kind of test kit detecting glycolated hemoglobin, described test kit includes:
First reagent, this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, precipitation total hemoglobin
Borate derivative；
Second reagent, this second reagent includes the buffering of the borate derivative for washing away uncombined glycolated hemoglobin
Liquid；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
Every liter of described first reagent includes following component:
Borate derivative 0.4g
Magnesium chloride 1.9g
Potassium chloride 22g
Barium chloride 6.2g
Glycyl amide hydrochloride 6.6g
Methanamide 25g
Potassium azide 8g
NPE 15g
Water surplus.
The structural formula of described borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 1 for carbon number.
Every liter of described second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 23g
Potassium azide 4g
Sodium chloride 0.4g
Methanamide 8g
Triton X-100 4g
Water surplus.
A kind of detection method using test kit detection glycolated hemoglobin described above for non-treatment purpose, including
Following steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, on
Under overturn 8 times, fully mix, stand 1min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 8s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 8s, chromatography device is placed on glycolated hemoglobin analysis and detects.
In described step A, every liter of first reagent includes following component:
Borate derivative 0.4g
Magnesium chloride 1.9g
Potassium chloride 22g
Barium chloride 6.2g
Glycyl amide hydrochloride 6.6g
Methanamide 25g
Potassium azide 8g
NPE 15g
Water surplus.
In described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 1 for carbon number.
In described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 23g
Potassium azide 4g
Sodium chloride 0.4g
Methanamide 8g
Triton X-100 4g
Water surplus.
Embodiment 2
A kind of test kit detecting glycolated hemoglobin, described test kit includes:
First reagent, this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, precipitation total hemoglobin
Borate derivative；
Second reagent, this second reagent includes the buffering of the borate derivative for washing away uncombined glycolated hemoglobin
Liquid；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
Every liter of described first reagent includes following component:
Borate derivative 0.45g
Magnesium chloride 2.2g
Potassium chloride 23g
Barium chloride 6.7g
Glycyl amide hydrochloride 7.2g
Methanamide 28g
Potassium azide 9g
NPE 18g
Water surplus.
The structural formula of described borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 2 for carbon number.
Every liter of described second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 25g
Potassium azide 4.5g
Sodium chloride 0.5g
Methanamide 9g
Triton X-100 4.5g
Water surplus.
A kind of detection method using test kit detection glycolated hemoglobin described above for non-treatment purpose, including
Following steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, on
Under overturn 9 times, fully mix, stand 1.5min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 9s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 9s, chromatography device is placed on glycolated hemoglobin analysis and detects.
In described step A, every liter of first reagent includes following component:
Borate derivative 0.45g
Magnesium chloride 2.2g
Potassium chloride 23g
Barium chloride 6.7g
Glycyl amide hydrochloride 7.2g
Methanamide 28g
Potassium azide 9g
NPE 18g
Water surplus.
In described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 2 for carbon number.
In described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 25g
Potassium azide 4.5g
Sodium chloride 0.5g
Methanamide 9g
Triton X-100 4.5g
Water surplus.
Embodiment 3
A kind of test kit detecting glycolated hemoglobin, described test kit includes:
First reagent, this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, precipitation total hemoglobin
Borate derivative；
Second reagent, this second reagent includes the buffering of the borate derivative for washing away uncombined glycolated hemoglobin
Liquid；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
Every liter of described first reagent includes following component:
Borate derivative 0.5g
Magnesium chloride 2.4g
Potassium chloride 24g
Barium chloride 7.2g
Glycyl amide hydrochloride 7.6g
Methanamide 30g
Potassium azide 10g
NPE 20g
Water surplus.
The structural formula of described borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 3 for carbon number.
Every liter of described second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 28g
Potassium azide 5g
Sodium chloride 0.6g
Methanamide 10g
Triton X-100 5g
Water surplus.
A kind of detection method using test kit detection glycolated hemoglobin described above for non-treatment purpose, including
Following steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, on
Under overturn 10 times, fully mix, stand 1-3min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 10s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 10s, chromatography device is placed on glycolated hemoglobin analysis and detects.
In described step A, every liter of first reagent includes following component:
Borate derivative 0.5g
Magnesium chloride 2.4g
Potassium chloride 24g
Barium chloride 7.2g
Glycyl amide hydrochloride 7.6g
Methanamide 30g
Potassium azide 10g
NPE 20g
Water surplus.
In described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 1-6 for carbon number.
In described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 28g
Potassium azide 5g
Sodium chloride 0.6g
Methanamide 10g
Triton X-100 5g
Water surplus.
Embodiment 4
A kind of test kit detecting glycolated hemoglobin, described test kit includes:
First reagent, this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, precipitation total hemoglobin
Borate derivative；
Second reagent, this second reagent includes the buffering of the borate derivative for washing away uncombined glycolated hemoglobin
Liquid；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
Every liter of described first reagent includes following component:
Borate derivative 0.55g
Magnesium chloride 2.6g
Potassium chloride 25g
Barium chloride 7.7g
Glycyl amide hydrochloride 8.1g
Methanamide 32g
Potassium azide 11g
NPE 22g
Water surplus.
The structural formula of described borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 4 for carbon number.
Every liter of described second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 30g
Potassium azide 5.5g
Sodium chloride 0.7g
Methanamide 11g
Triton X-100 5.5g
Water surplus.
A kind of detection method using test kit detection glycolated hemoglobin described above for non-treatment purpose, including
Following steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, on
Under overturn 8-12 time, fully mix, stand 1-3min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 8-12s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 8-12s, chromatography device is placed on glycolated hemoglobin analysis and detects.
In described step A, every liter of first reagent includes following component:
Borate derivative 0.55g
Magnesium chloride 2.6g
Potassium chloride 25g
Barium chloride 7.7g
Glycyl amide hydrochloride 8.1g
Methanamide 32g
Potassium azide 11g
NPE 22g
Water surplus.
In described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 4 for carbon number.
In described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 30g
Potassium azide 5.5g
Sodium chloride 0.7g
Methanamide 11g
Triton X-100 5.5g
Water surplus.
Embodiment 5
A kind of test kit detecting glycolated hemoglobin, described test kit includes:
First reagent, this first reagent includes for splitting erythrocyte, release glycolated hemoglobin, precipitation total hemoglobin
Borate derivative；
Second reagent, this second reagent includes the buffering of the borate derivative for washing away uncombined glycolated hemoglobin
Liquid；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
Every liter of described first reagent includes following component:
Borate derivative 0.6g
Magnesium chloride 2.9g
Potassium chloride 26g
Barium chloride 8.2g
Glycyl amide hydrochloride 8.6g
Methanamide 35g
Potassium azide 12g
NPE 25g
Water surplus.
The structural formula of described borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 5 for carbon number.
Every liter of described second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 33g
Potassium azide 6g
Sodium chloride 0.8g
Methanamide 12g
Triton X-100 6g
Water surplus.
A kind of detection method using test kit detection glycolated hemoglobin described above for non-treatment purpose, including
Following steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, on
Under overturn 12 times, fully mix, stand 1-3min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 12s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 12s, chromatography device is placed on glycolated hemoglobin analysis and detects.
In described step A, every liter of first reagent includes following component:
Borate derivative 0.6g
Magnesium chloride 2.9g
Potassium chloride 26g
Barium chloride 8.2g
Glycyl amide hydrochloride 8.6g
Methanamide 35g
Potassium azide 12g
NPE 25g
Water surplus.
In described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 5 for carbon number.
In described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 33g
Potassium azide 6g
Sodium chloride 0.8g
Methanamide 12g
Triton X-100 6g
Water surplus.
Use test kit and commercially available biochemical reagents, testing result such as table 1 institute of high performance liquid chromatography of embodiment 1-5
Show:
Table 1
| Testing result | Embodiment 1 | Embodiment 2 | Embodiment 3 | Embodiment 4 | Embodiment 5 |
| The test kit of the present invention | 3.6 | 5.7 | 6.8 | 9.3 | 11.2 |
| HPLC | 3.6 | 5.8 | 6.7 | 9.3 | 11.1 |
| Biochemical reagents | 3.3 | 5.4 | 6.2 | 9.7 | 10.5 |
As can be seen from the above table, the testing result of the present invention is close with the testing result of high performance liquid chromatography, and is better than
The testing result of commercially available biochemical reagents, highly sensitive, specificity good.The test kit of the present invention has only to the whole blood tip alive of trace
Blood sample, can realize the content of detection by quantitative glycolated hemoglobin in 2-3 minute, greatly improved the speed of examination, tool
Have highly sensitive, specificity good and advantage easy and simple to handle；Preparation method is simple, it is easy to large-scale production.
The detection method of the present invention has only to the whole blood peripheral blood sample alive of trace, can realize quantitatively in 2-3 minute
The content of detection glycolated hemoglobin, greatly improves the speed of examination, have highly sensitive, specificity good and easy and simple to handle
Advantage.
Above-described embodiment is the present invention preferably implementation, and in addition, the present invention can realize with alternate manner,
Without departing from obvious replacement any on the premise of present inventive concept all within protection scope of the present invention.
Claims (6)
1. the test kit detecting glycolated hemoglobin, it is characterised in that: described test kit includes:
First reagent, every liter of described first reagent includes following component:
Borate derivative 0.4-0.6g
Magnesium chloride 1.9-2.9g
Potassium chloride 22-26g
Barium chloride 6.2-8.2g
Glycyl amide hydrochloride 6.6-8.6g
Methanamide 25-35g
Potassium azide 8-12g
NPE 15-25g
Water surplus；
Second reagent, this second reagent includes the buffer of the borate derivative for washing away uncombined glycolated hemoglobin；
And chromatography device, this chromatography device includes the reaction film for retaining hemoglobin precipitation.
A kind of test kit detecting glycolated hemoglobin the most according to claim 1, it is characterised in that: described borate spreads out
Biological structural formula is:
,
Wherein, R1For azine, R2It is the alkyl of 1-6 for carbon number.
A kind of test kit detecting glycolated hemoglobin the most according to claim 1, it is characterised in that: every liter described second
Reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 23-33g
Potassium azide 4-6g
Sodium chloride 0.4-0.8g
Methanamide 8-12g
Triton X-100 4-6g
Water surplus.
4. one kind uses the test kit described in any one of claim 1-3 to detect HbAle egg for non-treatment and diagnostic purpose
White detection method, it is characterised in that: comprise the steps:
A, draw 5 μ L blood samples with capillary blood taking tube, be equipped with in the brown centrifuge tube of 200 μ L the first reagent, run up and down
Fall 8-12 time, fully mix, stand 1-3min at room temperature, obtain reactant liquor；
B, draw with pipettor 25 μ L above-mentioned reactant liquors dropping chromatography device in the middle of reaction film on；
After C, 8-12s, take on 25 μ L the second reagent dropwise reaction film in the middle of chromatography device；
After D, 8-12s, chromatography device is placed on glycolated hemoglobin analysis and detects.
A kind of detection method detecting glycolated hemoglobin for non-treatment and diagnostic purpose the most according to claim 4,
It is characterized in that: in described step A, the structural formula of borate derivative is:
,
Wherein, R1For azine, R2It is the alkyl of 1-6 for carbon number.
A kind of detection method detecting glycolated hemoglobin for non-treatment and diagnostic purpose the most according to claim 4,
It is characterized in that: in described step C, every liter of second reagent includes following component:
4-hydroxyethyl piperazine ethanesulfonic acid 23-33g
Potassium azide 4-6g
Sodium chloride 0.4-0.8g
Methanamide 8-12g
Triton X-100 4-6g
Water surplus.
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| CN105301261A (en) * | 2015-10-12 | 2016-02-03 | 广州江元医疗科技有限公司 | Kit for detecting HbAlc (Glycosylated Hemoglobin), and preparation method and using method of kit |
| CN105606418A (en) * | 2015-12-30 | 2016-05-25 | 李新华 | Integral kit detecting glycosylated hemoglobin and detection method thereof |
| US11474120B2 (en) | 2016-09-29 | 2022-10-18 | Green Cross Medical Science | Separable cassette for measuring glycated hemoglobin |
| EP3537156A4 (en) | 2016-11-04 | 2020-07-15 | Green Cross Medical Science | Measurement method for glycated hemoglobin ratio |
| CN107255724A (en) * | 2017-01-03 | 2017-10-17 | 深圳市惠安生物科技有限公司 | Real-time test reagent device of glycosylated hemoglobin and preparation method thereof |
| CN107153121B (en) * | 2017-04-12 | 2019-03-08 | 深圳市东邦生物医疗技术有限公司 | Glycosylated hemoglobin, hemoglobin detection kit and its detection method |
| CN107228940B (en) * | 2017-05-31 | 2019-09-24 | 吉林省汇酉生物技术股份有限公司 | A kind of detection reagent and method of cystatin C |
| CN112198320B (en) * | 2020-10-12 | 2023-12-22 | 青岛汉唐生物科技有限公司 | Glycosylated hemoglobin detection kit and detection method thereof |
| CN112198123B (en) * | 2020-10-12 | 2022-07-01 | 青岛汉唐生物科技有限公司 | Detection system and detection method for correcting influence of anemia on glycated hemoglobin measurement value |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5631364A (en) * | 1994-03-31 | 1997-05-20 | Axis Biochemicals Asa | Labelled boronic acid derivatives |
| CN1531654A (en) * | 2001-04-24 | 2004-09-22 | ������˹-ϣ���¹�˾ | Haemoglobin assay |
| CN102998463A (en) * | 2012-11-29 | 2013-03-27 | 英科新创(厦门)科技有限公司 | Method for measuring glycosylated hemoglobin and kit |
Family Cites Families (1)
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-
2015
- 2015-05-21 CN CN201510262100.6A patent/CN104897907B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5631364A (en) * | 1994-03-31 | 1997-05-20 | Axis Biochemicals Asa | Labelled boronic acid derivatives |
| CN1531654A (en) * | 2001-04-24 | 2004-09-22 | ������˹-ϣ���¹�˾ | Haemoglobin assay |
| CN102998463A (en) * | 2012-11-29 | 2013-03-27 | 英科新创(厦门)科技有限公司 | Method for measuring glycosylated hemoglobin and kit |
Non-Patent Citations (3)
| Title |
|---|
| Boronic acid functionalized superparamagnetic iron oxide nanoparticle as a novel tool for adsorption of sugar;S.Mohapatra et al.;《Materials Science and Engineering C》;20090831;第29卷(第7期);第2254-2260页 * |
| NycoCard HbAIc试剂盒测定糖化血红蛋白的临床应用;梁淑媛 等;《吉林医学》;20061231(第07期);第789页 * |
| 糖化血红蛋白快速定量检测试剂盒（挪威）型号：SA16NycoCard Reader Ⅱ-2;郑经理;《企博网》;20070831;第1-2页 * |
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